This project examines two aspects of the polyoma tumor antigens: (1) how structure is related to function within each protein and, (2) how the T antigens interact with cellular components to exert their regulatory effect on the cell. Recombinant DNA techniques are used to overproduce the T antigens and the individual proteins are purified from the resulting strains. Purified proteins can then tested for biological or biochemical function. Subsequently they can be used as reagents for raising banks of monoclonal antibodies for structural studies and as affinity reagents for studying interactios with host cell proteins. To complement these biochemical approaches, genetic studies will be done using retrovirus vectors which transduce the individual T antigens. A library of missense mutants of middle T antigens which have conditionally or absolutely lost transforming function will be made using the virus vectors. Interesting mutations will be characterized biochemically and sequenced at the DNA level to yield structure function data for the proteins. Together, the biochemical and genetic approaches will eventually allow us to overproduce mutant T-antigens so that they can be purified for further study. Such a situation should yield valuable information about the manner in which polyoma T antigens alter a cell to a transformed state.